Abstract

It has been commonly observed that gratuitous overexpression of proteins in Escherichia coli causes growth retardation. However, the molecular events involved in the metabolic response to the overexpression of proteins are still unclear. Here we used DNA microarray technology to characterize the changes in transcriptional patterns of selected host genes during protein overexpression. A nontoxic, soluble protein, LuxA (coded by luxA), which is the α-subunit of the luciferase heterodimer, was overexpressed for this purpose. A total of 132 E. coli genes, including those in the central metabolism, key biosynthetic pathways, and selected regulatory functions, were used as probes for detecting the level of mRNA transcripts in E. coli strains JM109, MC4100, and VJS676A during protein overexpression. Upon induction, these strains shared several common responses, such as the upregulation of glk and the heat shock genes as well as the downregulation of fba, ppc, atpA, and gnd. In addition, the biosynthesis genes glnA, glyA, and leuA were down-regulated in all three strains. Media-dependent responses were also observed in our study. For example, many respiratory genes that were upregulated in defined media showed an opposite effect in complex media under protein-overproducing conditions. These results demonstrate that gratuitous overexpression of proteins triggers a complex global response that involves several metabolic and regulatory systems. Explanations based on either existing knowledge of global regulations such as the heat shock response and the stringent response or stoichiometric analysis without regulatory considerations cannot account for the response induced by protein overexpression. (C) 2000 Academic Press.

abstract = "It has been commonly observed that gratuitous overexpression of proteins in Escherichia coli causes growth retardation. However, the molecular events involved in the metabolic response to the overexpression of proteins are still unclear. Here we used DNA microarray technology to characterize the changes in transcriptional patterns of selected host genes during protein overexpression. A nontoxic, soluble protein, LuxA (coded by luxA), which is the α-subunit of the luciferase heterodimer, was overexpressed for this purpose. A total of 132 E. coli genes, including those in the central metabolism, key biosynthetic pathways, and selected regulatory functions, were used as probes for detecting the level of mRNA transcripts in E. coli strains JM109, MC4100, and VJS676A during protein overexpression. Upon induction, these strains shared several common responses, such as the upregulation of glk and the heat shock genes as well as the downregulation of fba, ppc, atpA, and gnd. In addition, the biosynthesis genes glnA, glyA, and leuA were down-regulated in all three strains. Media-dependent responses were also observed in our study. For example, many respiratory genes that were upregulated in defined media showed an opposite effect in complex media under protein-overproducing conditions. These results demonstrate that gratuitous overexpression of proteins triggers a complex global response that involves several metabolic and regulatory systems. Explanations based on either existing knowledge of global regulations such as the heat shock response and the stringent response or stoichiometric analysis without regulatory considerations cannot account for the response induced by protein overexpression. (C) 2000 Academic Press.",

N2 - It has been commonly observed that gratuitous overexpression of proteins in Escherichia coli causes growth retardation. However, the molecular events involved in the metabolic response to the overexpression of proteins are still unclear. Here we used DNA microarray technology to characterize the changes in transcriptional patterns of selected host genes during protein overexpression. A nontoxic, soluble protein, LuxA (coded by luxA), which is the α-subunit of the luciferase heterodimer, was overexpressed for this purpose. A total of 132 E. coli genes, including those in the central metabolism, key biosynthetic pathways, and selected regulatory functions, were used as probes for detecting the level of mRNA transcripts in E. coli strains JM109, MC4100, and VJS676A during protein overexpression. Upon induction, these strains shared several common responses, such as the upregulation of glk and the heat shock genes as well as the downregulation of fba, ppc, atpA, and gnd. In addition, the biosynthesis genes glnA, glyA, and leuA were down-regulated in all three strains. Media-dependent responses were also observed in our study. For example, many respiratory genes that were upregulated in defined media showed an opposite effect in complex media under protein-overproducing conditions. These results demonstrate that gratuitous overexpression of proteins triggers a complex global response that involves several metabolic and regulatory systems. Explanations based on either existing knowledge of global regulations such as the heat shock response and the stringent response or stoichiometric analysis without regulatory considerations cannot account for the response induced by protein overexpression. (C) 2000 Academic Press.

AB - It has been commonly observed that gratuitous overexpression of proteins in Escherichia coli causes growth retardation. However, the molecular events involved in the metabolic response to the overexpression of proteins are still unclear. Here we used DNA microarray technology to characterize the changes in transcriptional patterns of selected host genes during protein overexpression. A nontoxic, soluble protein, LuxA (coded by luxA), which is the α-subunit of the luciferase heterodimer, was overexpressed for this purpose. A total of 132 E. coli genes, including those in the central metabolism, key biosynthetic pathways, and selected regulatory functions, were used as probes for detecting the level of mRNA transcripts in E. coli strains JM109, MC4100, and VJS676A during protein overexpression. Upon induction, these strains shared several common responses, such as the upregulation of glk and the heat shock genes as well as the downregulation of fba, ppc, atpA, and gnd. In addition, the biosynthesis genes glnA, glyA, and leuA were down-regulated in all three strains. Media-dependent responses were also observed in our study. For example, many respiratory genes that were upregulated in defined media showed an opposite effect in complex media under protein-overproducing conditions. These results demonstrate that gratuitous overexpression of proteins triggers a complex global response that involves several metabolic and regulatory systems. Explanations based on either existing knowledge of global regulations such as the heat shock response and the stringent response or stoichiometric analysis without regulatory considerations cannot account for the response induced by protein overexpression. (C) 2000 Academic Press.